As the plasminogen activator involved in fibrinolysis, known are tissue plasminogen activator (t-PA) produced by endothelial cells, and urokinase (UK), of which UK is conventionally well-known as a fibrinolysin. While UK has been normally purified from human urine or culture of human nephrocyte, production of UK by DNA recombination has been recently achieved (EP-A-154272). When used in a large amount, however, UK induces decomposition and activation of various factors inducing coagulation or fibrinolysis, and causes bleeding. The present inventors have already found that inactivated-type human urokinase precursor produced by human nephrocyte [EP-A-139447, J. Biol. Chem., 260, 12377 (1985)]permits, in contrast to UK, thrombolysis without bleeding [Cell Struc. Func., 10, 151 (1985)].
Human PUK has three domains, namely, epidermal growth factor (hereinafter abbreviated as EGF) domain, kringle domain, and enzyme activity domain [Hoppe-Seyler's Z. Physiol. Chem., 363, 1155 (1982)].
Incidentally, there has been heretofore attempted to modify human PUK in an effort to prolong half-life in blood. For example, there have been reported deletion of the entire EGF region (EP-A-253241), deletion of the first or the third loop in the EGF region (EP-A-398361), addition of a sugar chain by way of a specific partial structure obtained by replacing the amino acid in the EGF region (EP-A-398362), and so on.
On the other hand, while an attempt to increase affinity for fibrin can be evidenced by reports made of hybrid protein of UK or tPA with anti-fibrin antibody, and of incorporation of the kringle region of protein having high affinity for fibrin such as plasminogen into tPA or UK, there have been no reports made of an increased affinity for fibrin by an amino acid replacement.